Elizabeth A. Tovar, Curt J. Essenburg, Anderson Peck, L. Turner, Z. Madaj, Matthew C. P. Smith, J. Christensen, M. Melnik, E. Haura, Matthew R. Steensma, Carrie Graveel
{"title":"Abstract A48: MET and EGFR interaction promotes acquired resistance to kinase inhibition in TNBC","authors":"Elizabeth A. Tovar, Curt J. Essenburg, Anderson Peck, L. Turner, Z. Madaj, Matthew C. P. Smith, J. Christensen, M. Melnik, E. Haura, Matthew R. Steensma, Carrie Graveel","doi":"10.1158/1557-3125.ADVBC17-A48","DOIUrl":null,"url":null,"abstract":"Triple-negative breast cancer (TNBC) has the worst prognosis of all breast cancers, and the molecular heterogeneity within TNBC heightens the challenge of developing effective targeted therapies. Receptor tyrosine kinases (RTKs), in particular MET and EGFR, are promising therapeutic targets for TNBC due to their high expression in multiple molecular TNBC subtypes and the tendency for cancers to become “kinase addicted.” We and others have demonstrated that MET is highly expressed in TNBC and its expression correlates with poor prognosis. EGFR expression is also elevated in up to 72% of TNBCs and correlates with poor prognosis in TNBC patients. Recently, we demonstrated that MET and EGFR are coordinately and highly expressed across all TNBC subtypes and the efficacy of combined MET and EGFR inhibition in TNBC patient-derived xenograft (PDX) models. Even though MET and EGFR receptors are actionable targets due to their high activity in TNBC, crosstalk between MET and EGFR has been implicated in therapeutic resistance to kinase inhibitors in several cancer types and needs to be evaluated in TNBC. There is strong evidence demonstrating the critical role of redundant RTK signaling networks in resistance to tyrosine kinase inhibitors (TKIs). Specifically, crosstalk between MET and EGFR has been implicated in therapeutic resistance to EGFR or MET inhibitors in colon, gastric, and lung cancers. Since MET and EGFR are highly expressed in a substantial proportion of TNBCs, we used TNBC PDX models and TNBC cell lines to interrogate mechanisms of resistance to MET and EGFR kinase inhibition. We hypothesized that coexpression, interaction, and activation of MET and EGFR promote acquired TKI resistance and an adaptive kinome response in TNBC. Using TNBC PDX models that highly express both MET and EGFR, we evaluated MET and EGFR activity, localization, and interaction after 3 days or 3 weeks of treatment with MET (glesatinib, crizotinib) and/or EGFR inhibitors (erlotinib). After 3 weeks of dual MET and EGFR inhibition in TNBC PDXs, only a small population of “resistant” cells remained. In these resistant populations, we observed increased MET and EGFR colocalization and increased MET and EGFR activation. To understand how colocalization of MET and EGFR promotes signaling redundancy and crosstalk, we asked whether MET and EGFR directly interact in TNBC PDX using a proximity ligation assay (PLA). PLA uses antibody specificity to detect direct protein-protein interactions at physiologic levels in vitro and in vivo. We observed a significant increase in MET EGFR interactions only in resistant cell populations that had been treated with both MET and EGFR inhibitors. AKT and ERK signaling were also increased in TKI-resistant cells, indicating that these are key survival pathways in resistance. Interestingly, we did not observe an increase in MET-EGFR activity or interaction in PDX tumors treated for only 3 days. Even though TNBC cells (HCC70) showed increased colocalization of MET and EGFR with combined glesatinib and erlotinib treatment, there was not a significant increase in MET-EGFR interactions. Evaluation of MET-EGFR interaction in primary TNBC patient samples revealed significant MET-EGFR interactions in a MET-amplified tumor and lymph node metastases. To our knowledge, this is the first time MET-EGFR interactions have been observed in TNBC in vivo and in human patients. Moreover, MET-EGFR interactions are able to maintain MET, EGFR, ERK, and AKT activity through a yet-unknown mechanism. These results imply that MET EGFR interaction may be a unique mechanism of resistance to kinase inhibition that may be inherent in human TNBCs with genomic amplification of MET or EGFR. Citation Format: Elizabeth Tovar, Curt Essenburg, Anderson Peck, Lisa Turner, Zachary Madaj, Matthew Smith, James Christensen, Marianne Melnik, Eric Haura, Matthew Steensma, Carrie Graveel. MET and EGFR interaction promotes acquired resistance to kinase inhibition in TNBC [abstract]. In: Proceedings of the AACR Special Conference: Advances in Breast Cancer Research; 2017 Oct 7-10; Hollywood, CA. Philadelphia (PA): AACR; Mol Cancer Res 2018;16(8_Suppl):Abstract nr A48.","PeriodicalId":20897,"journal":{"name":"Resistance Mechanisms","volume":"52 1","pages":""},"PeriodicalIF":0.0000,"publicationDate":"2018-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Resistance Mechanisms","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1158/1557-3125.ADVBC17-A48","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0
Abstract
Triple-negative breast cancer (TNBC) has the worst prognosis of all breast cancers, and the molecular heterogeneity within TNBC heightens the challenge of developing effective targeted therapies. Receptor tyrosine kinases (RTKs), in particular MET and EGFR, are promising therapeutic targets for TNBC due to their high expression in multiple molecular TNBC subtypes and the tendency for cancers to become “kinase addicted.” We and others have demonstrated that MET is highly expressed in TNBC and its expression correlates with poor prognosis. EGFR expression is also elevated in up to 72% of TNBCs and correlates with poor prognosis in TNBC patients. Recently, we demonstrated that MET and EGFR are coordinately and highly expressed across all TNBC subtypes and the efficacy of combined MET and EGFR inhibition in TNBC patient-derived xenograft (PDX) models. Even though MET and EGFR receptors are actionable targets due to their high activity in TNBC, crosstalk between MET and EGFR has been implicated in therapeutic resistance to kinase inhibitors in several cancer types and needs to be evaluated in TNBC. There is strong evidence demonstrating the critical role of redundant RTK signaling networks in resistance to tyrosine kinase inhibitors (TKIs). Specifically, crosstalk between MET and EGFR has been implicated in therapeutic resistance to EGFR or MET inhibitors in colon, gastric, and lung cancers. Since MET and EGFR are highly expressed in a substantial proportion of TNBCs, we used TNBC PDX models and TNBC cell lines to interrogate mechanisms of resistance to MET and EGFR kinase inhibition. We hypothesized that coexpression, interaction, and activation of MET and EGFR promote acquired TKI resistance and an adaptive kinome response in TNBC. Using TNBC PDX models that highly express both MET and EGFR, we evaluated MET and EGFR activity, localization, and interaction after 3 days or 3 weeks of treatment with MET (glesatinib, crizotinib) and/or EGFR inhibitors (erlotinib). After 3 weeks of dual MET and EGFR inhibition in TNBC PDXs, only a small population of “resistant” cells remained. In these resistant populations, we observed increased MET and EGFR colocalization and increased MET and EGFR activation. To understand how colocalization of MET and EGFR promotes signaling redundancy and crosstalk, we asked whether MET and EGFR directly interact in TNBC PDX using a proximity ligation assay (PLA). PLA uses antibody specificity to detect direct protein-protein interactions at physiologic levels in vitro and in vivo. We observed a significant increase in MET EGFR interactions only in resistant cell populations that had been treated with both MET and EGFR inhibitors. AKT and ERK signaling were also increased in TKI-resistant cells, indicating that these are key survival pathways in resistance. Interestingly, we did not observe an increase in MET-EGFR activity or interaction in PDX tumors treated for only 3 days. Even though TNBC cells (HCC70) showed increased colocalization of MET and EGFR with combined glesatinib and erlotinib treatment, there was not a significant increase in MET-EGFR interactions. Evaluation of MET-EGFR interaction in primary TNBC patient samples revealed significant MET-EGFR interactions in a MET-amplified tumor and lymph node metastases. To our knowledge, this is the first time MET-EGFR interactions have been observed in TNBC in vivo and in human patients. Moreover, MET-EGFR interactions are able to maintain MET, EGFR, ERK, and AKT activity through a yet-unknown mechanism. These results imply that MET EGFR interaction may be a unique mechanism of resistance to kinase inhibition that may be inherent in human TNBCs with genomic amplification of MET or EGFR. Citation Format: Elizabeth Tovar, Curt Essenburg, Anderson Peck, Lisa Turner, Zachary Madaj, Matthew Smith, James Christensen, Marianne Melnik, Eric Haura, Matthew Steensma, Carrie Graveel. MET and EGFR interaction promotes acquired resistance to kinase inhibition in TNBC [abstract]. In: Proceedings of the AACR Special Conference: Advances in Breast Cancer Research; 2017 Oct 7-10; Hollywood, CA. Philadelphia (PA): AACR; Mol Cancer Res 2018;16(8_Suppl):Abstract nr A48.
三阴性乳腺癌(TNBC)是所有乳腺癌中预后最差的,TNBC内部的分子异质性增加了开发有效靶向治疗的挑战。受体酪氨酸激酶(rtk),特别是MET和EGFR,由于它们在多种分子TNBC亚型中的高表达和癌症成为“激酶依赖”的趋势,是TNBC有希望的治疗靶点。我们和其他人已经证明MET在TNBC中高表达,其表达与不良预后相关。EGFR表达在高达72%的TNBC患者中升高,并与TNBC患者预后不良相关。最近,我们证明MET和EGFR在所有TNBC亚型中是协调和高表达的,并且在TNBC患者来源的异种移植(PDX)模型中,MET和EGFR联合抑制的效果。尽管MET和EGFR受体因其在TNBC中的高活性而成为可操作的靶点,但MET和EGFR之间的串扰与几种癌症类型对激酶抑制剂的治疗抗性有关,需要在TNBC中进行评估。有强有力的证据表明,冗余的RTK信号网络在对酪氨酸激酶抑制剂(TKIs)的抗性中起关键作用。具体来说,MET和EGFR之间的串扰与结肠癌、胃癌和肺癌对EGFR或MET抑制剂的治疗性耐药有关。由于MET和EGFR在相当大比例的TNBC中高度表达,我们使用TNBC PDX模型和TNBC细胞系来探究MET和EGFR激酶抑制的抗性机制。我们假设MET和EGFR的共表达、相互作用和激活促进了TNBC中获得性TKI抗性和适应性激酶反应。使用高表达MET和EGFR的TNBC PDX模型,我们评估了MET和EGFR活性、定位和在MET(格莱沙替尼、克里唑替尼)和/或EGFR抑制剂(厄洛替尼)治疗3天或3周后的相互作用。在TNBC pdx中进行MET和EGFR双重抑制3周后,只有一小部分“耐药”细胞仍然存在。在这些耐药人群中,我们观察到MET和EGFR共定位增加,MET和EGFR激活增加。为了了解MET和EGFR的共定位如何促进信号冗余和串扰,我们使用近端结联试验(PLA)研究MET和EGFR是否在TNBC PDX中直接相互作用。PLA利用抗体特异性检测体外和体内生理水平上的直接蛋白相互作用。我们观察到MET和EGFR抑制剂治疗的耐药细胞群中MET和EGFR相互作用的显著增加。AKT和ERK信号也在tki耐药细胞中升高,表明它们是耐药的关键生存途径。有趣的是,我们没有观察到MET-EGFR活性的增加或PDX肿瘤治疗仅3天的相互作用。尽管TNBC细胞(HCC70)在格沙替尼和厄洛替尼联合治疗后MET和EGFR共定位增加,MET和EGFR相互作用并未显著增加。对原发性TNBC患者样本中MET-EGFR相互作用的评估显示,met扩增的肿瘤和淋巴结转移中存在显著的MET-EGFR相互作用。据我们所知,这是首次在TNBC体内和人类患者中观察到MET-EGFR相互作用。此外,MET-EGFR相互作用能够通过一种未知的机制维持MET、EGFR、ERK和AKT的活性。这些结果表明MET或EGFR相互作用可能是一种独特的抵抗激酶抑制的机制,这种机制可能是具有MET或EGFR基因组扩增的人类tnbc固有的。引文格式:Elizabeth Tovar, Curt Essenburg, Anderson Peck, Lisa Turner, Zachary Madaj, Matthew Smith, James Christensen, Marianne Melnik, Eric Haura, Matthew Steensma, Carrie Graveel。MET和EGFR相互作用促进TNBC对激酶抑制的获得性抗性[摘要]。摘自:AACR特别会议论文集:乳腺癌研究进展;2017年10月7-10日;费城(PA): AACR;中华肿瘤杂志,2018;16(8):1 - 8。